Fig 2.

CpoS variants with mutations in coiled-coil motifs display differential abilities to counteract cell-autonomous defenses. (A) Predicted domain structure of CpoS(L2) expressed by wild-type, mutant, and complemented strains [N-terminal (N), transmembrane (TM), intra-inclusion (I), and C-terminal (C) domains; coiled-coil (CC) motifs; FLAG (F) tag]. Topology and CC motifs were predicted using Phobius tool (25) and Paircoil2 (26), respectively. Disruptive effects of mutations on CC motifs were predicted using both Paircoil2 (26) and Coils (27). (B) Relative expression levels of CpoS variants in infected HeLa cells (10 IFU/cell, 32 hpi) determined by western blot analysis using anti-FLAG antibodies. (C) Localization of CpoS variants in infected HeLa cells (5 IFU/cell, 22 hpi, scale = 20 µm). (D–G) Ability of CpoS variants to prevent cell death (D), block ISRE-driven luciferase expression (E), block STING translocation (F), and restore IFU (i.e., EB) production (G). Details as described in Fig. 1D through F. Note, the experiments were conducted together with those shown in Fig. 1D through F and share the same controls. All quantitative results in Fig. 2 represent mean ± SD [n = 6 (A2EN in panel D), n = 3 (else); one-way ANOVA: indicated are significant differences compared to pCpoS(L2) (B) or CTL2 (else, unless indicated otherwise)].